论文部分内容阅读
目的构建人胰岛素样生长因子1(IGF-1)分泌型真核表达载体,并检测表达产物的生物活性。方法从人肝细胞克隆IGF-1基因,并将其连入Psec Tag/FRT/V5-His载体,用脂质体法转染CHO细胞,将转染细胞上清培养骨髓基质干细胞,用MTT法测定骨髓基质干细胞的增殖情况。结果成功扩增210bp的IGF-1基因,表达产物经SDS-PAGE分析及Western bolt检测具有7.7KD特异性条带。骨髓基质干细胞的增殖随转染细胞上清的浓度、转染细胞上清培养时间的增加而增加。结论成功构建分泌型IGF-1真核表达载体,其表达产物对骨髓基质干细胞具有促增殖作用。
Objective To construct the secreting eukaryotic expression vector of human insulin-like growth factor 1 (IGF-1) and to detect the biological activity of the expressed product. Methods The IGF-1 gene was cloned from human hepatocytes and transfected into Psec Tag / FRT / V5-His vector. CHO cells were transfected by lipofectamine. The bone marrow stromal cells were cultured in supernatant of transfected cells. MTT assay The proliferation of bone marrow stromal stem cells was measured. Results The 210bp IGF-1 gene was successfully amplified. The expressed product was identified by SDS-PAGE and Western blot with a specific band of 7.7 kD. The proliferation of bone marrow stromal stem cells increased with the concentration of transfected cell supernatant and the culture time of transfected cell supernatant increased. Conclusion The eukaryotic expression vector of secreting IGF-1 was successfully constructed, and its expression product could promote the proliferation of bone marrow stromal stem cells.